Detection and identification of Vibrio parahaemolyticus by multiplex PCR and DNA-DNA hybridization on a microarray |
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| Authors: | Rongzhi Wang Jiadong Huang Wei Zhang Guangmei Lin Junwei Lian Libin Jiang Hongcong Lin Songfa Wang Shihua Wang |
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| Affiliation: | [1]The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China [2]College of Chemistry and Chemical Engineering, University ofJinan, Jinan 250022, China |
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| Abstract: | In this paper,we developed a rapid and accurate method for the detection of Vibrio parahaemolyticus strains,using multiplex PCR and DNA-DNA hybridization.Multiplex PCR was used to simultaneously amplify three diagnostic genes(tlh,tdh and fia)that serve as molecular markers of V.parahaemolyticus.Biotinylated PCR products were hybridized to primers immobilized on a microarray,and detected by chemiluminesce with avidin-conjugated alkaline phosphatase.With this method,forty-five samples were tested.Eight known virulent strains (tlh+/tdh+/fia+)and four known avirulent strains(tlh+/tdh-/fla+)of the V.parahaemolyttcus were successtuny aetectea,ana no non-spectnc hybridization and cross-hybridization reaction were found from fifteen closely-related strains(tin-/tdh-/fta+)or the Vibrio spp.In addition,all the other eighteen strains of non-Vibrio bacteria(tlh-/tdh-/fla-)gave negative results.The DNA microarray successfully distinguished V.parahaemolyticus from other Vibrio spp.The results demonstrated that this was an efficient and robust method for identifying virulent strains of V.parahaemolyticus. |
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| Keywords: | Vtbrio parahaemolyticus Multiplex PCR Hybridization Gene microarray |
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